Dear Label_Name,

Greetings! I hope you are well.

I am writing on behalf of an international publishing house, Science and Education Publishing, USA (http://www.sciepub.com).

Science and Education Publishing (SciEP) is a multidisciplinary open access publisher of Journals covering the fields of Science, Technology and Medicine. All SciEP publications are subject to high-quality peer review, editorial and production processes and are freely available online.

Welcome to the April, 2013 edition of SciEP newsletter, providing monthly summaries of articles on science & education related topics of interest to scientist and professionals. The mission of our journals is to provide an open access forum for publishing new findings on issues pertinent to scientific and academic field. We also invite you to submit original research papers, review articles, short reports, or case studies are considered for publication in our Multidisciplinary Journals.

American Journal of Public Health Research
DOI: 10.12691/ajphr-1-1-3

The Implementation of a Tele-Homecare System with Service Oriented Architecture and HL7 Message Transmission Standard
Ching-Sung Wang

American Journal of Medical and Biological Research
DOI: 10.12691/ajmbr-1-1-2

The Influence of Ethanol on Pyruvate Kinases Activity in Vivo, in Vitro, in Silico
Lelevich Sergey Vladimirovich, Khrustalev Vladislav Victorovich, Barkovsky Eugene Victorovich, Shedogubova Tatyana Aleksandrovna

American Journal of Medical Sciences and Medicine
DOI: 10.12691/ajmsm-1-1-1

Clinical and Histological Features in Wegener Granulomatosis
Li Cavoli Gioacchino, Passantino Rita, Bono Luisa, Tortorici Calogera, Ferrantelli Angelo, Giammarresi Carlo, Rotolo Ugo

American Journal of Infectious Diseases and Microbiology
DOI: 10.12691/ajidm-1-1-2

The Curse of Lymphatic Filariasis: Would the Continual Use of Diethylcarbamazine Eliminate this Scourge in Papua New Guinea?
Anne Lanham, Lillian Mwanri

We have released issues of some journals on February 2013.
American Journal of Public Health Research
American Journal of Medical Sciences and Medicine
American Journal of Medical and Biological Research
American Journal of Pharmacological Sciences
American Journal of Clinical Medicine Research
American Journal of Nursing Research
American Journal of Cancer Prevention
American Journal of Medicine Studies
American Journal of Infectious Diseases and Microbiology
American Journal of Cardiovascular Disease Research
American Journal of Biomedical Research
American Journal of Mechanical Engineering
American Journal of Microbiological Research
Journal of Preventive Medicine
American Journal of Food and Nutrition
American Journal of Hypertension Research
American Journal of Sports Science and Medicine
Journal of Cancer Research and Treatment
American Journal of Zoological Research
Journal of Biomedical Engineering and Technology
American Journal of Microbiological Research
Biomedical Science and Engineering

We are pleased to announce that submissions are now being accepted for peer review for all the journals across all our subject areas in SciEP.

Please submit your manuscript via our online submission system:
http://www.sciepub.com/author/signin

To unsubscribe from our mailing list, please click on the following link: unsubscribe

I am looking forward to hearing from you.

Kind regards,

David Rain
Email:join@sciepub.com
Science and Education Publishing (http://www.sciepub.com)

Label_Title

Dear Label_Name,

Greetings! I hope you are well.

I am writing on behalf of an international publishing house, Science and Education Publishing, USA (http://www.sciepub.com).

Science and Education Publishing (SciEP) is a multidisciplinary open access publisher of Journals covering the fields of Science, Technology and Medicine. All SciEP publications are subject to high-quality peer review, editorial and production processes and are freely available online.

Welcome to the April, 2013 edition of SciEP newsletter, providing monthly summaries of articles on science & education related topics of interest to scientist and professionals. The mission of our journals is to provide an open access forum for publishing new findings on issues pertinent to scientific and academic field. We also invite you to submit original research papers, review articles, short reports, or case studies are considered for publication in our Multidisciplinary Journals.

American Journal of Public Health Research
DOI: 10.12691/ajphr-1-1-3

The Implementation of a Tele-Homecare System with Service Oriented Architecture and HL7 Message Transmission Standard
Ching-Sung Wang

American Journal of Medical and Biological Research
DOI: 10.12691/ajmbr-1-1-2

The Influence of Ethanol on Pyruvate Kinases Activity in Vivo, in Vitro, in Silico
Lelevich Sergey Vladimirovich, Khrustalev Vladislav Victorovich, Barkovsky Eugene Victorovich, Shedogubova Tatyana Aleksandrovna

American Journal of Medical Sciences and Medicine
DOI: 10.12691/ajmsm-1-1-1

Clinical and Histological Features in Wegener Granulomatosis
Li Cavoli Gioacchino, Passantino Rita, Bono Luisa, Tortorici Calogera, Ferrantelli Angelo, Giammarresi Carlo, Rotolo Ugo

American Journal of Infectious Diseases and Microbiology
DOI: 10.12691/ajidm-1-1-2

The Curse of Lymphatic Filariasis: Would the Continual Use of Diethylcarbamazine Eliminate this Scourge in Papua New Guinea?
Anne Lanham, Lillian Mwanri

We have released issues of some journals on February 2013.
American Journal of Public Health Research
American Journal of Medical Sciences and Medicine
American Journal of Medical and Biological Research
American Journal of Pharmacological Sciences
American Journal of Clinical Medicine Research
American Journal of Nursing Research
American Journal of Cancer Prevention
American Journal of Medicine Studies
American Journal of Infectious Diseases and Microbiology
American Journal of Cardiovascular Disease Research
American Journal of Biomedical Research
American Journal of Mechanical Engineering
American Journal of Microbiological Research
Journal of Preventive Medicine
American Journal of Food and Nutrition
American Journal of Hypertension Research
American Journal of Sports Science and Medicine
Journal of Cancer Research and Treatment
American Journal of Zoological Research
Journal of Biomedical Engineering and Technology
American Journal of Microbiological Research
Biomedical Science and Engineering

We are pleased to announce that submissions are now being accepted for peer review for all the journals across all our subject areas in SciEP.

Please submit your manuscript via our online submission system:
http://www.sciepub.com/author/signin

To unsubscribe from our mailing list, please click on the following link: unsubscribe

I am looking forward to hearing from you.

Kind regards,

David Rain
Email:join@sciepub.com
Science and Education Publishing (http://www.sciepub.com)

Ten Times Normal Anti Tissue Transglutaminase Level Detected By Enzyme Linked Immunosorbent Assay (ELISA), But Normal Level by Chemiluminescent Assay (CLIA)

Fatemeh Elham Mahjoub*, Mehrnaz Farasat, Mehri Najafi Sani, Azizollah Yousefi, Pegah Pakzadian, Fatemeh Farahmand, Farzaneh Moatamed, Pejman Rohani, Shahindoqht Homayoun Valiane, Alireza Nourbakhsh, Touran Akbari

Tehran University of Medical Sciences, Tehran, Iran, Associate Professor in Pathology-Roshan Azma Pathobiology Laboratory, Tehran, Iran

Abstract: According to the new guidelines of the European Society for Pediatric Gastroenterology Hepatology and Nutrition [1], it is mandatory to test for anti tissue transglutaminase IgA (anti TTG IgA) and serum IgA in first step in the diagnosis of celiac disease. A diagnosis of celiac disease can be made when the anti-TTG IgA titer is ten times normal (with normal Ig A level) and anti-endomysial antibody (EMA) is positive in the second blood sample, without requiring biopsy. In this article, we present cases of anti TTG IgA levels more than ten times the upper limit of normal determined by ELISA (enzyme-linked immunosorbent assay). In some patients, EMA was checked and in all of these cases upper endoscopy and duodenal biopsy was performed for confirmation, Although biopsy was performed from the bulb and the second part of the duodenum, and the number of biopsies were adequate; no evidence of celiac disease was observed on histological examination. These patients were recruited, and another blood sample was taken and anti TTG IgA was checked by a chemiluminescence immunoassay (CLIA) and the levels were reported normal.

Keywords: Celiac Disease, Serology, Anti Tissue Transglutaminase, Enzyme Linked Immunosorbent Assay (ELISA), Chemiluminescent Immunoassay (CLIA)

Received June 05, 2025; Revised July 07, 2025; Accepted July 14, 2025

Citation: Fatemeh Elham Mahjoub, Mehrnaz Farasat, Mehri Najafi Sani, Azizollah Yousefi, Pegah Pakzadian, Fatemeh Farahmand, Farzaneh Moatamed, Pejman Rohani, Shahindoqht Homayoun Valiane, Alireza Nourbakhsh, Touran Akbari, Ten Times Normal Anti Tissue Transglutaminase Level Detected By Enzyme Linked Immunosorbent Assay (ELISA), But Normal Level by Chemiluminescent Assay (CLIA). International Journal of Celiac Disease, 13 (1). 1-3. January 2025. doi:10.12691/ijcd-13-1-1

1. Introduction

Celiac disease is a lifelong autoimmune condition requiring strict avoidance of gluten containing foods.

In low- and middle-income countries, bread is a major food item for families, including their children. It is hard to share information with families that their child is not able to eat bread with other family members. So the diagnosis of celiac disease should be declared after finding strong evidence.

Based on ESPAGHAN 2020, eligible children for no biopsy approach for celiac disease diagnosis are those with high serum IgA class antibody concentration against transglutaminase 2 values (more than 10 times the upper limit of normal) with appropriate tests and positive endomysial antibodies (EMA-IgA) in a second serum sample.

The method for evaluating anti TTG IgA level is not mentioned in the guidelines and it is only mentioned that only calibration curve – based immunoassays should be used. Although most western countries have greater access to standardized and high tech instruments for the evaluation of anti TTG IgA level, however the method mostly used in our country is ELISA.

Endoscopic studies are frequently performed in our country, especially in the pediatric age group, either following the suspicion of a pediatric gastro - enterologist or because the family insists on doing so. We encountered cases in which elevated anti TTG IgA level (ten times higher than normal) were reported in recent years, in whom a biopsy was taken from the bulb and duodenum and the sample was sent to our laboratory.

Later, we checked the anti TTG IgA level using a chemiluminescent assay, and the level was reported as normal.

This has led to confusion among most clinicians and raised anger in families about faults and flaws in laboratory tests.

Here, we discuss the matter just to mention that in some cases, ten times normal anti TTG IgA level by ELISA method maybe false and the level should be confirmed by CLIA method on a second blood sample.

2. Materials and Methods

Between 2018 and March 2024, we encountered 6 children with elevated anti TTG IgA levels ten times normal, they all underwent upper endoscopy and biopsy.

Anti TTG IgA levels were measured using ELISA in different laboratories (some are the best in our country) and with different types of kits. EMA was measured on second blood sample in some of the patients and some had negative result.

In all of these cases, biopsy was performed due to the insistence of the parents or in some cases due to suspicion of the pediatric gastro-enterologist.

At least four biopsy samples obtained from the bulb and distal duodenum in each patient (except patient number 4: one piece from bulb and two pieces from D2) and sample was sent to our laboratory (Roshan Azma Pathobiology Laboratory, Tehran, Iran). New Marsh grading system was used for histopathology report [2].

Our patients approve that the information can be used in scientific processes before submitting the sample. Besides this is a retrospective study and cases presented here are anonymous and personal information is well preserved.

After processing and embedding, two slides were taken from each sample and stained with Hematoxylin & Eosin.

The slides were observed by a pathologist with approximately 27 years of experience in pediatric gastrointestinal pathology.

In all cases, another blood sample was analyzed using the chemiluminescence method (Liaison Instruments) [3].

It is important to state that none of these children were on gluten free diet at the time of upper endoscopy and biopsy.

3. Results

Six patients with anti TTG IgA antibody levels more than 10 times the upper limit of normal detected.

Four patients were male and the median age was 8.4 years (5-11.5 years). The demographic details are depicted in attached table (Table 1).

EMA was checked and in patients number 5 and 6, it was positive (other patients had negative EMA level)

Two patients underwent repeat ELISA tests on a second blood sample in another laboratory and the results were the same (patients number 1 and number 5).

All of the above mentioned cases had normal histology and normal anti TTG IgA levels by chemiluminescence method. In addition, we had patients with elevated anti TTG IgA with ELISA who had normal histology, but patients did not referred for assessment of anti TTG IgA level with chemiluminescent method and were not included in this study.

Despite the highly elevated anti TTG IgA levels detected by ELISA, these cases were regarded as false positive and no gluten free diet was administered. Seven other patients had anti TTG IgA levels between 5 and 10 times normal. All patients had normal histology. The second blood sample was checked by the chemiluminescent method and the level was reported as normal.

Table 1. Demographic Characteristics And Serologic Markers of Our Patients

4. Discussion

Tissue transglutaminase (TTG or TG2) is a 78-kDa, calcium-dependent enzyme belonging to the glutamyl -transferases family [4,5]. TTG acts as an autoantigen in celiac disease [6]. Diagnostic Serological tests for anti TTG antibodies have been claimed to have strong sensitivity (99%) and specificity (> 90%) for identifying celiac disease and has superseded other older tests, such as the anti gliadin antibody test [7].

Modern anti-TTG assays use a recombinant human protein as antigen [8].

Based on a recent study there is no significant differences in diagnostic accuracy among ELISA, CLIA, and fluorometric enzyme-linked immunoassay (FEIA) for detecting anti-TTG Ig A, but while different methods exist, ELISA remains highly accurate [9].

Based on ESPGHAN guidelines only antibody tests with proper calibrator curve-based calculation should be used [1].

Although, the ESPGHAN guideline recommends that celiac disease can be diagnosed without biopsy in cases with anti TTG IgA levels ten times above normal and if EMA is positive in second blood sample, we had cases in which the assessment was performed using ELISA, but the biopsy was normal, and later, anti TTG IgA levels were reported to be normal by the CLIA method.

This could be due to the effect of sanctions on the supplementation of kits in Iran. Surprisingly, there have been cases in which anti TTG IgA levels were reported to be very high in the above mentioned laboratories and in biopsy, celiac disease was approved.

Therefore, the problem seems to be a random error that cannot be traced to a specific laboratory and/or kit.

In two of our patients (number 5 and 6) even EMA was positive however the clinician wanted to confirm celiac disease by biopsy and upper endoscopy was performed despite ESPGHAN guidelines. In patient number 5 first EMA was done by ELISA method and was positive but another sample was checked with immunofluorescent method and was reported as negative. In patient number 6, EMA was checked by ELISA and was highly elevated but test was repeated on another blood sample in a different laboratory and was reported as negative. This shows that ELISA method for assaying of EMA is not flawless.

We believe that in countries such as Iran, where anti TTG IgA is mostly checked by ELISA, it is prudent to check anti TTG IgA in the second sample by another method (in two of our cases, the second sample was checked by ELISA in another laboratory and the result was the same) or to perform biopsies, especially when there is any suspicion of the diagnosis of celiac disease.

The main ingredient in the diet of Iranians is bread and gluten- free products are usually very expensive.

In this regard accurate diagnosis is very important.

Unfortunately access to chemiluminescence or fluorescent immunoassays is difficult due to sanctions and rare laboratories, such as the Sophia Pathobiology Laboratory, perform the test using the Liaison instrument in Iran [3].

However, this laboratory has many difficulties maintaining the supply of instruments and kits in Iran.

Pediatric gastro - enterologists in Iran are willing to perform upper endoscopy for all patients with elevated levels of anti TTG IgA despite the ESPGHAN guidelines due to mistrust of test results. Sometimes they put the patient on a gulten free diet despite normal biopsy findings and interpret it as a hidden type of celiac disease.

However, we believe that in cases where anti TTG IgA levels are reported to be normal by the CLIA method, it is useless to suffer families by this laboratory fault, and it is prudent not to recommend gluten free diet for these kids.

The above findings may lead to further studies comparing ELISA with other methods for the assessment of anti TTG antibodies, and we hope that the test method will be mentioned in upcoming guidelines.

References

[1]   | 2020 Update of the Guidelines for diagnosing Coeliac disease published by the European Society of Paediatric Gastroenterology, Hepatology and Nutrition. 2020-07-02.
[2]   | Oberhuber G, Histopathology of celiac disease. , August 2000, Pages 368-372.
[3]   | https://int.diasorin.com/en/immunodiagnostics/gastrointestinal-diseases/liaison-ttg-iga.
[4]   | Király R, Demény M, Fésüs L. "Protein transamidation by transglutaminase 2 in cells: a disputed Ca2+-dependent action of a multifunctional protein". The FEBS Journal. December 2011; 278 (24): 4717–39.
[5]   | Klöck C, Diraimondo TR, Khosla C.). "Role of transglutaminase 2 in celiac disease pathogenesis". Seminars in Immunopathology. July 2012; 34 (4): 513–22. PMID 22437759.
[6]   | Di Sabatino A, Vanoli A, Giuffrida P, Luinetti O, Solcia E, Corazza GR.. "The function of tissue transglutaminase in celiac disease". Autoimmunity Reviews. August 2012; 11 (10): 746–53.

[7]  
Anil K. Verma, Simona Gatti, Elena Lionetti, Tiziana Galeazzi, Chiara Monachesi, Elisa Franceschini, Linda Balanzoni, Novella Scattolo,k Mauro Cinquetti, and Carlo Catassi. Comparison of Diagnostic Performance of the IgA Anti-tTG Test vs IgA Anti–Native Gliadin Antibodies Test in Detection of Celiac Disease in the General Population. Clinical Gastroenterology and Hepatology. 2018; 16: 1997–1998.

[8]   | Sblattero D, Berti I, Trevisiol C, Marzari R, Tommasini A, Bradbury A, Fasano A, Ventura A, Not T. "Human recombinant tissue transglutaminase ELISA: an innovative diagnostic assay for celiac disease". The American Journal of Gastroenterology. May 2000; 95 (5): 1253–7.
[9]   | Pjetraj, D.; Pulvirenti, A.; Moretti, M.; Gatti, S.; Catassi, G.N.; Catassi, C.; Lionetti, E. Diagnostic Accuracy of IgA Anti-Transglutaminase Assessed by Chemiluminescence: A Systematic Review and Meta-Analysis. Nutrients 2024, 16, 2427.